The studies in this proposal address the specialized functions of cholangiocytes, the epithelial cells that line the lumen of bile ducts and contribute to bile formation. CI-channels play a key role in the regulation of secretion, though the cellular mechanisms involved are largely unknown. Recent findings from our laboratory suggest a novel pathway, purinergic signaling, as a potent determinant of secretion. It is proposed that extracellular ATP functions as an autocrine/paracrine factor to regulate cholangiocyte secretion. The Specific Aims are to i) determine the pathways mediating regulated ATP release in biliary epithelia, ii) evaluate the intracellular signals linking purinergic receptor binding to channel activation and cholangiocyte secretion, and iii) determine the specific channel(s) involved in mediating cholangiocyte secretion in response to extracellular ATP. The long-term goal of these studies is to define the cellular mechanisms involved in cholangiocyte secretion and bile formation. The findings are directly relevant to a broad range of cholestatic liver diseases characterized by impaired cholangiocyte function. Defining the cellular mechanisms responsible for the regulation of biliary secretion will serve as a basis for the development of therapeutic interventions to modulate bile formation for the treatment of cholestatic liver disorders. The proposed studies emphasize complementary measurements of transport from single ion channels, intact biliary cells, and novel models of intact biliary epithelium that retain secretory polarity. An integrative approach combining electrophysiology, molecular biology, and fluorescence imaging will be applied to the proposed studies. Studies will be performed at the University of Colorado Health Sciences Center under the preceptorship of Dr. Gregory Fitz. During this mentored research period new techniques in advanced patch clamp analysis and molecular biology will be developed. The long-term goals of the applicant encompass two complimentary objectives, i) to form a fundamental understanding of biliary physiology to serve as a basis for the development of effective treatments of cholestatic liver disorders, and ii) to develop a independent and successful research career. The Mentored Clinical Scientist Development Award will provide the protected time, resources, and mentored environment to allow the applicant to achieve these goals.